MPEA – makes sensitive miRNA labeling easy

febit’s new protocol Microfluidic Primer Extension Assay (MPEA) offers an easy way to gain highly sensitive labeling of miRNA for the Geniom® Microarray Technology: The MPEA combines the specificity of a hybridization assay with the discriminatory power of an enzymatic extension.

How does this work?

MPEA uses unlabeled miRNAs for hybridization on highly flexible microfluidic microarrays. In a second step, the Klenow fragment of DNA polymerase I is added directly into the channels of the microfluidic chip, where the specific elongation of the bound miRNAs takes place.

The sensitivity is high enough to analyze nanogram amounts of total RNA coming from formalin fix, paraffin embedded (FFPE) tissue samples, Fine Needle Aspiration or Laser Capture Microdissection without amplification.

MPEA offers you:

• Easy-to-use protocol for Geniom RT Analyzer
• Higher sensitivity and specificity
• High discriminatory power
• Very low mismatch ratio
• Low amount of sample RNA needed

For more detailed information about the advantages and the practice of the microfluidic-based enzymatic on-chip labeling MPEA please read one of these publications:

• Small Molecules with Big Impact in Cancer
• Impact of MicroRNAs in Cancer Research

Comparison of two microarray-based assays used for miRNA profiling. In both cases, labeling was done using biotinylated nucleotides, which enables the subsequent staining with a streptavidin-phycoerythrin-conjugate.
A): For the conventional hybridization assay, miRNAs are labeled first and hybridized afterwards to the microarray containing reverse complementary oligonucleotides.
B): Principle of the MPEA: The hybridized, unlabeled miRNA functions as a primer for an enzymatic elongation in which biotinylated nucleotides are incorporated.